Takimoto M, Tomonaga T, Matunis M, Avigan M, Krutzsch H, Dreyfuss G, Levens D
Laboratory of Pathology, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892.
J Biol Chem. 1993 Aug 25;268(24):18249-58.
A homopurine/homopyrimidine-like sequence is found 100-150 base pairs upstream of the human c-myc promoter P1. This element, termed the CT-element, has been shown to augment expression from P1, and it serves as a positive transcriptional element when coupled to a heterologous promoter in vivo and in vitro. Synthetic oligonucleotides comprising this element were used to form DNA-protein complexes in electrophoretic mobility shift assays. By using conventional and affinity methods, 61- and 34-kDa proteins were shown to be associated with these complexes. Amino acid sequence analysis and immunological methods have identified these proteins as heterogeneous ribonucleoprotein particle (hnRNP) proteins K and A1. Surprisingly, hnRNP protein K binds to the pyrimidine-rich strand of the CT-element in a sequence-specific manner as well as to the double-stranded molecule. Cotransfection of vectors encoding hnRNP protein K in the sense or anti-sense orientations with reporter plasmids driven by wild-type or mutant CT-elements demonstrates that hnRNP protein K augments gene expression in a cis-element-dependent manner. Taken together, these results suggest that hnRNP protein K may play a role in the transcriptional regulation of the human c-myc gene.
在人类c-myc启动子P1上游100 - 150个碱基对处发现了一个类同嘌呤/同嘧啶序列。这个元件，称为CT元件，已被证明可增强P1的表达，并且在体内和体外与异源启动子偶联时，它作为一个正向转录元件发挥作用。在电泳迁移率变动分析中，使用包含该元件的合成寡核苷酸来形成DNA - 蛋白质复合物。通过使用常规方法和亲和方法，显示61 kDa和34 kDa的蛋白质与这些复合物相关联。氨基酸序列分析和免疫学方法已将这些蛋白质鉴定为不均一核糖核蛋白颗粒（hnRNP）蛋白K和A1。令人惊讶的是，hnRNP蛋白K以序列特异性方式结合到CT元件富含嘧啶的链以及双链分子上。将编码有义或反义方向hnRNP蛋白K的载体与由野生型或突变型CT元件驱动的报告质粒共转染表明，hnRNP蛋白K以顺式元件依赖的方式增强基因表达。综上所述，这些结果表明hnRNP蛋白K可能在人类c-myc基因的转录调控中发挥作用。
